Inactivation of negative regulators such as Cdk inhibitors and activation of positive regulators such as cyclins serve to promote inappropriate proliferation. Although direct genetic mechanisms are clearly important in determining the status of the cell cycle regulatory apparatus, it is now clear that alterations in the levels of gene products through post-transcriptional mechanisms can also contribute in important ways to proliferative disturbances. Alterations in the balance of growth regulators often reflect changes in the rates of protein turnover via the ubiquitinproteasome system. Components of the ubiquitin system have been linked to decreased levels of p53 and the Cdk inhibitor p27, and increased levels of oncoproteins such as cyclins, c-myc, and beta-catenin in human cancer. We have found that the oncoprotein Skp2, an E3 ubiquitin ligase, is induced in prostate cancer and PIN lesions, and this inversely correlates with decreased levels of its target p27 and positively correlates with recurrence. This proposal seeks to further understand how ubiquitin-mediated proteolysis contributes to prostate cancer through four specific aims. First, we will address the role of Skp2 in promoting proliferation and transformation of prostate cells using in vitro and in vivo model systems. Second, components of the ubiquitination pathway are frequently altered in cancer. We will use genomic approaches to identify components of the ubiquitination system that are altered in prostate cancer. In addition, current evidence implicates a network of interacting protein kinases and ubiquitin ligases in the regulation of a number of growth promoting genes. We will establish the status of these pathways in normal prostate and prostate cancer. Third, we will correlate alterations in the status of ubiquitination components and their targets with clinical and pathological parameters. Fourth, a neoadjuvant clinical trial will be performed to elucidate the impact of the proteasome inhibitor PS341 on the constellation of growth promoting and inhibiting proteins that are controlled through the ubiquitin-proteasome pathway.